Movement of macromolecules between the nucleus and the cytoplasm is a highly orchestrated process essential for key cellular functions such as protein synthesis and gene regulation. Transport in and out of the nucleus begins with recognition at the transported cargo by a receptor. After docking at the nuclear pore, the cargo-receptor complex somehow moves through the aqueous pore channel, which is lined with nucleoporins. Loading and unloading of cargoes and interaction of cargo- receptor complexes with the nuclear pore are defining events of nuclear transport. How these events actually occur is poorly understood and remains one of the major unanswered questions in cell biology. The proposed research focuses on the transport of macromolecules in and out of the nucleus. The experiments employ the genetically tractable yeast S. cerevisiae and combine genetic, biochemical and cell biological approaches. The overall goals of the proposal are to determine how: 1) certain cargoes are recognized by their nuclear transport receptor; 2) receptors together with their cargo move through the nuclear pore channel; and 3) certain factors mediate only nuclear protein export. Significance of the research has basic and applied components. Key nuclear transport are highly conserved, making studies with model eukaryotes relevant to the process in all cells. From a more applied angle, components of the nuclear transport machinery are implicated in several cancers. For example, gene fusions with nucleoporins have been found in at least two types of leukemias. In one case, the fusion protein leads to mislocalization of the nucleoporin inside the nucleus. In addition, one nuclear transport receptor is amplified in some breast cancers and has been suggested to play a role in resistance to TNF- induced apoptotic death. Finally, many viruses, such as HIV, exploit the endogenous nuclear import and export machinery to propagate. HIV Rev interacts with the nuclear export receptor and Vpr with the nuclear import receptor and certain nucleoporins. Theses interactions are required for HIV infection of non-dividing macrophages. Understanding the process of nuclear transport and how it is regulated, may lead to new anti-viral drug targets and ways to enhance viral-based gene therapies.